Petit M A, Dervyn E, Rose M, Entian K D, McGovern S, Ehrlich S D, Bruand C
Laboratoire de Génétique Microbienne, INRA, Jouy en Josas, France.
Mol Microbiol. 1998 Jul;29(1):261-73. doi: 10.1046/j.1365-2958.1998.00927.x.
The only DNA helicase essential for Escherichia coli viability is DnaB, the chromosome replication for helicase. In contrast, in Bacillus subtilis, in addition to the DnaB counterpart called DnaC, we have found a second essential DNA helicase, called PcrA. It is 40% identical to the Rep and UvrD DNA helicases of E. coli and 61% identical to the PcrA helicase of Staphylococcus aureus. This gene is located at 55 degree on the chromosome and belongs to a putative operon together with a ligase gene (lig) and two unknown genes named pcrB and yerH. As PcrA was essential for cell viability, conditional mutants were constructed. In such mutants, chromosomal DNA synthesis was slightly decreased upon PcrA depletion, and rolling-circle replication of the plasmid pT181 was inhibited. Analysis of the replication intermediates showed that leading-strand synthesis of pT181 was prevented upon PcrA depletion. To compare PcrA with Rep and UvrD directly, the protein was produced in rep and uvrD mutants of E. coli. PcrA suppressed the UV sensitivity defect at a uvrD mutant but not its mutator phenotype. Furthermore, it conferred a Rep-phenotype on E. coli. Altogether, these results show that PcrA is an helicase used for plasmid rolling-circle replication and suggest that it is also involved in UV repair.
对大肠杆菌生存能力至关重要的唯一DNA解旋酶是DnaB,即用于解旋酶的染色体复制酶。相比之下,在枯草芽孢杆菌中,除了名为DnaC的DnaB对应物外,我们还发现了第二种必需的DNA解旋酶,称为PcrA。它与大肠杆菌的Rep和UvrD DNA解旋酶有40%的同一性,与金黄色葡萄球菌的PcrA解旋酶有61%的同一性。该基因位于染色体的55度处,与一个连接酶基因(lig)以及两个名为pcrB和yerH的未知基因属于一个假定的操纵子。由于PcrA对细胞生存能力至关重要,因此构建了条件突变体。在这类突变体中,PcrA缺失时染色体DNA合成略有减少,质粒pT181的滚环复制受到抑制。对复制中间体的分析表明,PcrA缺失时pT181的前导链合成被阻止。为了直接将PcrA与Rep和UvrD进行比较,该蛋白在大肠杆菌的rep和uvrD突变体中产生。PcrA抑制了uvrD突变体的紫外线敏感性缺陷,但没有抑制其诱变表型。此外,它赋予大肠杆菌一种Rep表型。总之,这些结果表明PcrA是一种用于质粒滚环复制的解旋酶,并表明它也参与紫外线修复。